We have determined the antigenic profile of natural killer (NK) cells from C57BL/6 mice that have lytic activity to RLol tumor cells, in a 5-hr 51Cr release assay. To perform the antigenic typing more effectively, we enriched the NK populations by using discontinuous BSA density gradient and established a correlation between NK activity and Nk-1 antigen in each layer. As NK activity and Nk-1 antigen are both criteria for measuring NK cells, we determined the antigenic role of NK cells by monitoring both the change in NK activity and Nk-1 plus cells after elimination of cells with various antisera to lymphocytic alloantigens. We have found that all NK cells expressed Nk-1, Oa-4, and Oa-5 antigens; about 50 percent of NK cells expressed Thy-1 and Oa-2 antigens, and 25 percent of NK cells also expressed Lyt-1 antigen. NK cells did not express significant amount of Oa-1, Oa-3, Lyt-2, and Lyb-2 antigens. Therefore, spontaneous NK cells are heterogeneous with regard to their surface antigens, some of which are T cell antigens. However, the present antigenic profile does not allow us to classify NK cells in the conventional T cell lineage.